Chronic lymphocytic leukemia (CLL) is the most prevalent leukemia in the United States with almost 5,000 attributable deaths per year. Despite significant advances in the understanding of the biology of CLL, it remains incurable and the cause of CLL is unknown. A more complete understanding of the biology of pre-emergent CLL will lead to novel prevention and treatment strategies. Monoclonal B cell lymphocytosis (MBL) is an asymptomatic hematologic syndrome characterized by small accumulations of clonal B lymphocytes in the peripheral blood. Most MBL share phenotypic characteristics with CLL: CD5+, CD19+, CD20dim, CD23+, and sIgdim. The biologic characteristics and clinical implications of MBL remain unclear. Our central hypothesis is that MBL are antigen-driven, post-germinal center CD5+ B cell expansions with variable potential for progression to CLL. To date, we have identified 73 MBL cases from 430 individuals ascertained from familial CLL kindreds both at Duke University and Genetic Epidemiology of CLL (GEC) consortium sites. In our preliminary investigations, we have found that MBL commonly have mutated immunoglobulin heavy chains (IgVH), express lower levels of surface IgM than CLL, and show enhanced B cell receptor (BCR) signaling when compared to memory B cells. To further explore these observations, three complementary specific aims are proposed: (i) to perform a detailed cellular and molecular biological analysis of MBL cases including single cell immunoglobulin gene analysis and detailed immunophenotyping; (ii) to explore BCR dependent signaling pathways in CLL and MBL and (iii) to follow a cohort of family-associated MBL subjects longitudinally to evaluate for evolution of the MBL clone, development of CLL, and other clinical outcomes. To investigate the biologic characteristics of MBL, we will perform detailed immunophenotyping and fluorescence-activated cell sorting (FACS) to purify MBL cells from MBL subjects identified through screening of Duke and GEC CLL families. We propose detailed evaluation of the B cell receptor to evaluate antigenic exposure and investigate whether receptor revision is a mechanism of by which IgVH unmutated MBL and CLL arises. BCR dependent signaling in MBL and CLL will be evaluated to determine the key mediators of the malignant phenotype in CLL; these signaling pathways will be targeted in vitro for CLL cell cytotoxicity. Finally, longitudinal evaluation to identify ongoing antigen refinement in MBL will clarify whether MBL are quiescent, good risk CLL or if MBL are dynamic populations that acquire additional somatic mutations as part of a step-wise transformation to CLL. In summary, MBL provides an important window into the biology of pre-emergent CLL. Careful investigation of subjects with MBL will illuminate the transforming events that lead to CLL, the role of antigenic stimulation in early CLL leukemogenesis, the signaling pathways that favor disease progression, and allow for MBL risk stratification for development of CLL.